The reverse transcriptase (RT) of Type 1 Human Immunodeficiency Virus (HIV-1) plays an indispensable role in the life cycle of the virus. Its premier function is the synthesis of a double-stranded DNA copy of the RNA genome for integration into the host chromosome. This is achieved by the concerted application of a number of innate activities including minus-strand DNA synthesis via an RNA-dependent DNA polymerase activity, concomitant degradation of the template RNA strand via an RNase H activity, and plus-strand DNA synthesis via a DNA-dependent DNA polymerase activity (Baltimore, D. (1970) Nature 226:1209; Temin, H. M. and Mizutani, S. (1970) Nature 226:1211; Gilboa, E. et al. (1979) Cell 18:93-100; Goff, S. P. (1990) J. Acq. Imm. Defic. Syndr. 3:93-100; Peliska, J. A. and Benkovic, S. J. (1992) Science 258:1112-1118). Because the cells HIV-1 infects contain no endogenous RT, it must also possess a mechanism to ensure its packaging into the mature viral particle to guarantee its presence in the succeeding infection.
HIV-1 is generally accepted as the etiological agent of Acquired Immune Deficiency Syndrome (AIDS). The importance of its function in the life cycle of HIV-1 and the lack of a natural function in the host cell make RT a preferred target for antiviral agents.
Several types of HIV-1 RT inhibitors are known. Many, such as AZT (3'-azido-2',3'-dideoxythymidine), are nucleoside analogs, which when incorporated into polynucleotides by HIV-1 RT, result in chain termination. (Kedar, P. S. et al. (1990) Biochem, 29:3603-3611; Huang, P. et al. (1990) J. Biol. Chem. 265:11914-11918). Other nucleoside analogs that inhibit HIV-1 RT include ddC (2',3'-dideoxycytidine) and ddI (2',3'-dideoxyinosine) Inhibitors that are not nucleoside analogs have also been described. These include dipyridodiazepinones (e.g., Merluzzi, V. J. et al. (1990) Science 250:1411-1413; Kopp, E. B. et al. (1991) Nuc. Acids Res. 19(11):3035- 3039), tetrahydro-imidazo 4,5,1-jk! 1,4!-benzodiazepin-2(1H)-one and -thione (TIBO) derivatives (e.g., Pauwels, T. et al. (1990) Nature 343:470-474), and catechin derivatives (e.g., Nakane, H., and Ono, K. (1990) Biochem. 29:2841-2845). These nonnucleosides inhibit by mechanisms other than direct competition for substrate binding sites (Kopp, E. B. et al. (1991) Nuc. Acids Res. 19(11): 3035-3039).
A family of phosphorodithioate-linked ssDNA nucleotides have been described with the property of inhibiting HIV-1 RT activity at K.sub.i values ranging from 0.5-180 nM (Marshall and Caruthers, (1993), Science 259:1564-1570). The specific sequences of these nucleotides were based on the sequence of various nucleic acid substrates of HIV-RT.
RNA pseudoknots that bind specifically to the polymerase active site of HIV-1 RT and inhibit the RNA-dependent DNA polymerase activity have already been identified using SELEX (U.S. patent application Ser. No. 07/964,624, which is specifically incorporated herein by reference; Tuerk, C. et al. (1992) Proc. Natl. Acad. Sci., U.S.A. 89:6988-6992).
The development of high affinity DNA ligands capable of inhibiting HIV-1 reverse transcriptase would be useful in the treatment of Type 1 Human Immunodeficiency Virus. Herein described are high affinity ssDNA ligand inhibitors of HIV-1 reverse transcriptase.